The present invention is directed to a process for the diagnosis of long QT syndrome (LQT). LQT has been associated with specific genes including HERG, SCN5A, KVLQT1 and KCNE1. LQT may be hereditary and due to specific mutations in the above genes or it may be acquired, e.g., as a result of treatment with drugs given to treat cardiac arrhythmias or of treatment with other types of medications such as antihistamines or antibiotics such as erythromycin. The acquired form of LQT is the more prevalent form of the disorder. It had previously been shown that the HERG gene encodes a K.sup.+ channel which is involved in the acquired form of LQT. It is shown that increasing the K.sup.+ levels in patients taking drugs to prevent cardiac arrhythmias may decrease the chances of the acquired form of LQT from developing and can be used as a preventive measure. Also, this knowledge can now be used to develop drugs which may activate this K.sup.+ channel and which could be given in conjunction with the drugs presently used to treat cardiac arrhythmias. Activation of the K.sup.+ channel should decrease the risk of developing LQT and torsade de pointes.
The publications and other materials used herein to illuminate the background of the invention or provide additional details respecting the practice, are incorporated by reference, and for convenience are respectively grouped in the appended List of References.
Although sudden death from cardiac arrhythmias is thought to account for 11% of all natural deaths, the mechanisms underlying arrhythmias are poorly understood (Kannel, 1987; Willich et al., 1987). One form of long QT syndrome (LQT) is an inherited cardiac arrhythmia that causes abrupt loss of consciousness, syncope, seizures and sudden death from ventricular tachyarrhythmias, specifically torsade de pointes and ventricular fibrillation (Ward, 1964; Romano, 1965; Schwartz et al., 1975; Moss et al., 1991). This disorder usually occurs in young, otherwise healthy individuals (Ward, 1964; Romano, 1965; Schwartz, 1975). Most LQT gene carriers manifest prolongation of the QT interval on electrocardiograms, a sign of abnormal cardiac repolarization (Vincent et al., 1992). The clinical features of LQT result from episodic cardiac arrhythmias, specifically torsade de pointes, named for the characteristic undulating nature of the electrocardiogram in this arrhythmia. Torsade de pointes may degenerate into ventricular fibrillation, a particularly lethal arrhythmia. Although LQT is not a common diagnosis, ventricular arrhythmias are very common; more than 300,000 United States citizens die suddenly every year (Kannel et al., 1987; Willich et al., 1987) and, in many cases, the underlying mechanism may be aberrant cardiac repolarization. LQT, therefore, provides a unique opportunity to study life-threatening cardiac arrhythmias at the molecular level. A more common form of this disorder is called "acquired LQT" and it can be induced by many different factors, particularly treatment with certain medications and reduced serum K.sup.+ levels (hypokalemia).
Autosomal dominant and autosomal recessive forms of the hereditary form of this disorder have been reported. Autosomal recessive LQT (also known as Jervell-Lange-Nielsen syndrome) has been associated with congenital neural deafness; this form of LQT is rare (Jervell and Lange-Nielsen, 1957). Autosomal dominant LQT (Romano-Ward syndrome) is more common, and is not associated with other phenotypic abnormalities. A disorder very similar to inherited LQT can also be acquired, usually as a result of pharmacologic therapy (Schwartz et al., 1975; Zipes, 1987).
In 1991, the complete linkage between autosomal dominant LQT and a polymorphism at HRAS was reported (Keating et al., 1991a; Keating et al., 1991b). This discovery localized LQT1 to chromosome 11p15.5 and made presymptomatic diagnosis possible in some families. Autosomal dominant LQT was previously thought to be genetically homogeneous, and the first seven families that were studied were linked to 11p15.5 (Keating et al., 1991b). In 1993, it was found that there was locus heterogeneity for LQT (Benhorin et al., 1993; Curran et al., 1993b; Towbin et al., 1994). Two additional LQT loci were subsequently identified, LQT2 on chromosome 7q35-36 (nine families) and LQT3 on 3p21-24 (three families) (Jiang et al., 1994). The genes responsible for LQT at these loci were subsequently identified. These are KVLQT1 (LQT1), HERG (LQT2), and SCN5A (LQT3) (Wang et al., 1996; Curran et al., 1995; Wang et al., 1995; U.S. Pat. No. 5,599,673). Later, KCNE1 (LQT5) was also associated with long QT syndrome (Splawski et al., 1997; Duggal et al., 1998). These genes encode ion channels involved in generation of the cardiac action potential. Mutations can lead to channel dysfunction and delayed myocellular repolarization. Because of regional heterogeneity of channel expression within the myocardium, the aberrant cardiac repolarization creates a substrate for arrhythmia. KVLQT1 and KCNE1 are also expressed in the inner ear (Neyroud et al., 1997; Vetter et al., 1996). It has been demonstrated that homozygous or compound heterozygous mutations in each of these genes can cause deafness and the severe cardiac phenotype of the Jervell and Lange-Nielsen syndrome (Neyroud et al., 1997; Splawski et al., 1997; Schultze-Bahr et al., 1997; Tyson et al., 1997). Loss of functional channels in the ear apparently disrupts the production of endolymph, leading to deafness. Several families remain unlinked to the known loci, indicating additional locus heterogeneity for LQT. This degree of heterogeneity suggests that distinct LQT genes may encode proteins that interact to modulate cardiac repolarization and arrhythmia risk.
Presymptomatic diagnosis of LQT is currently based on prolongation of the QT interval on electrocardiograms. QTc (QT interval corrected for heart rate) greater than 0.44 second has traditionally classified an individual as affected. Most LQT patients, however, are young, otherwise healthy individuals, who do not have electrocardiograms. Moreover, genetic studies have shown that QTc is neither sensitive nor specific (Vincent et al., 1992). The spectrum of QTc intervals for gene carriers and non-carriers overlaps, leading to misclassifications. Non-carriers can have prolonged QTc intervals and be diagnosed as affected. Conversely, some LQT gene carriers have QTc intervals of .ltoreq.0.44 second but are still at increased risk for arrhythmia. Correct presymptomatic diagnosis is important for effective, gene-specific treatment of LQT.
Genetic screening using mutational analysis can improve presymptomatic diagnosis. The presence of a mutation would unequivocally distinguish affected individuals and identify the gene underlying LQT even in small families and sporadic cases. To facilitate the identification of LQT-associated mutations, we defined the genomic structure of HERG and designed primer pairs for the amplification of each exon. Single strand conformational polymorphism (SSCP) analyses identified additional mutations in HERG.
In 1994, Warmke and Ganetzky identified a novel human cDNA, human ether a-go-go related gene (HERG, Warnke and Ganetzky, 1994). HERG was localized to human chromosome 7 by PCR analysis of a somatic cell hybrid panel (Warnke and Ganetzky, 1994). The function of the protein encoded by HERG was not known, but it has predicted amino acid sequence homology to potassium channels. HERG was isolated from a hippocampal cDNA library by homology to the Drosophila ether a-go-go gene (eag), which encodes a calcium-modulated potassium channel (Bruggemann et al., 1993). HERG is not the human homolog of eag, however, sharing only .about.50% amino acid sequence homology. The function of HERG was unknown, but it was strongly expressed in the heart and was hypothesized to play an important role in repolarization of cardiac action potentials and was linked to LQT (Curran et al., 1995).
Acquired LQT usually results from therapy with medications that block cardiac K.sup.+ channels (Roden, 1988). The medications most commonly associated with LQT are antiarrhythmic drugs (e.g., quinidine, sotalol) that block the cardiac rapidly-activating delayed rectifier K.sup.+ current, I.sub.Kr, as part of their spectrum of pharmacologic activity. Other drugs may also cause acquired LQT. These include antihistamines and some antibiotics such as erythromycin. I.sub.Kr has been characterized in isolated cardiac myocytes (Balser et al., 1990; Follmer et al., 1992; Sanguinetti and Jurkiewicz, 1990; Shibasaki, 1987; T. Yang et al., 1994), and is known to have an important role in initiating repolarization of action potentials.
To define the physiologic role of HERG, the full-length cDNA was cloned and the channel was expressed in Xenopus oocytes. Voltage-clamp analyses of the resulting currents revealed that HERG encodes a K.sup.+ channel with biophysical characteristics nearly identical to I.sub.Kr. These data suggest that HERG encodes the major subunit for the I.sub.Kr channel, and provide a mechanistic link between some forms of inherited and drug-induced LQT.